The development of rapid, sensitive, simple to use, and inexpensive biosensors is of significant continuing interest to applications ranging from medical diagnostics to biowarfare agent detection. Over the past five years, a technique has been developed termed “Arrayed Imaging Reflectometry”, or AIR, that relies on the initial creation of a near-null reflectance condition in a thin film on the surface of a reflective substrate. When illuminated with s-polarized light at a wavelength and angle tuned to the material system used, perturbation of this near-null reflectance condition due to target binding is readily observed as an increase in the reflected intensity. This allows for parallel detection of analytes with high sensitivity and dynamic range. AIR employs an exceptionally simple detection system, containing a single-wavelength light source such as a helium-neon laser (an ideal wavelength for AIR chips based on silicon/silicon dioxide), a set of optics to polarize, expand, and collimate the beam, a fixed sample stage, and a CCD camera. To date, AIR has been employed for the detection of oligonucleotides (Lu et al., “Reflective Interferometric Detection of Label-free Oligonucleotides,” Anal Chem 76:4416-4420 (2004)), proteins derived from pathogenic E. coli (Mace et al., “A Theoretical and Experimental Analysis of Arrayed Imaging Reflectometry as a Sensitive Proteomics Technique,” Anal Chem 78:5578-5583 (2006)) as well as O15:H-E. coli itself (Horner et al., “A Proteomic Biosensor for Enteropathogenic E. Coli,” Biosensors and Bioelectronics 21:1659-1663 (2006)), and human cytokines (Mace et al., “Detection of Human Proteins Using Arrayed Imaging Reflectometry,” Biosensors and Bioelectronics 24:334-337 (2008)). The latter also allowed the demonstration of the suitability of employing AIR for detection of proteins in human serum, a challenging milieu for any label-free biodetection system.
Direct detection of pathogen-derived proteins or nucleic acids is obviously of crucial importance for the development of diagnostics. However, one can also detect the traces of the human body's exposure to a pathogen through the immune response: specific antibodies to pathogen-derived antigens are potentially useful as an indicator of exposure, and of immunity. So-called “antigen arrays” have been employed for a wide range of diseases, and recent examples include organisms responsible for leprosy (Duthie et al., “Selection of Antigens and Development of Prototype Tests for Point-of-Care Leprosy Diagnosis,” Clin Vaccine Immunol 15:1590-1597 (2008)), Lyme disease (Xu et al., “Profiling the Humoral Immune Response to Borrelia burgdorferi Infection with Protein Microarrays,” Microbial Pathogenesis 45:403-407 (2008), bovine tuberculosis (Whelan et al., “Multiplex Immunoassay for Serological Diagnosis of Myobacterium bovis Infection in Cattle,” Clin Vaccine Immunol 15:1834-1838 (2008), and Q fever (Beare et al., “Candidate Antigens for Q Fever Serodiagnosis Revealed by Immunoscreening of a Coxiella burnetti Protein Microarray,” Clin Vaccine Immunol 15:1771-1779 (2008)).
Detection of soluble proteins is fundamental to many aspects of immunology, but it is still a challenge for one method to achieve high sensitivity, robustness with biological fluids, multiplexing of many related analytes, in a rapid detection method with low costs. Array-based optical techniques for specific protein detection have the potential to address all of these objectives. Using specific capture antibodies, it is possible to detect very low concentrations of proteins with high specificity in a very rapid assay. Another major advantage is that no labeled detection reagents are needed, i.e., the analyte is detected directly.
It would be desirable to provide an array of immobilized antigens that can be used to screen for antibodies against infectious agents and vaccines involving multiple similar specificities, e.g., distinguishing between different strains of an infectious agent such as a virus, based on the immune response generated by these infectious agents.
The present invention is directed to overcoming this and other deficiencies in the art.